Bacterial Etiology of Wound Infection and Antibiotic Susceptibility Pattern of the Isolates

1. Presentation and Viva-voce examination on
Bacterial Etiology of Wound Infection and Antibiotic
Susceptibility Pattern of the Isolates
A
Dissertation
Submitted to the Department of Microbiology
Kantipur College of Medical Sciences
(Affiliated to Tribhuvan University)
In Partial Fulfillment of the Requirements for the Award of the
Degree of Master of Science in Microbiology (Medical)
By
Shreejeet Shrestha
Department of Microbiology
Kantipur College of Medical Sciences
(Affiliated to Tribhuvan University)
Sitapaila, Kathmandu, Nepal
2010

2. Introduction
• The presence of microorganisms in a wound is not unusual but not all
wounds support the same range and number of species, the outcome of
wound infection depends on interaction of complex host and microbial
factors.
• Since wound colonization is most frequently polymicrobial involving
numerous microorganisms that are potentially pathogenic, any wound is at
some risk of becoming infected.
• Wound infections may occur as a result of penetrating trauma from plants,
animals, guns, knives or other various objects and are mainly caused by
overcrowding, lack of general cleanliness, poor socioeconomic condition
and lack of education too.
• Wound infections are one of the most common hospital acquired infections
and are an important cause of morbidity and account for 70-80% mortality.
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3. Introduction
• Wound infections can be caused by different groups of microorganisms
like bacteria, fungi and protozoa. However, different microorganisms exist
in polymicrobial communities especially in the margins of wounds and in
chronic wounds
• Most commonly isolated aerobic microorganisms include Staphylococcus
aureus, Coagulase-negative staphylococci (CoNS), Enterococci,
Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumonia, Proteus
mirabilis, Acinetobacter etc
• Infected wounds may not yield pathogens by culture owing to the
fastidious nature of some pathogens, or if the patients has received an
antimicrobial therapy
• The widespread uses of antibiotics, together with length of time over which
they have been available have led to major problems of resistance
pathogens contributing to morbidity and mortality.
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4. Objective
• General Objective
 To study the bacterial etiology of wound infection and their antibiotic
susceptibility pattern among the patients visiting B and B Hospital.
• Specific Objectives
 To isolate and identify the bacterial pathogens from patients with
wound infections.
 To describe the distribution pattern of bacterial pathogens in relation to
age and sex of the patients.
 To describe the seasonal distribution of bacterial pathogens among the
patients.
 To study the antibiotic susceptibility pattern of the isolated bacterial
pathogen.
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5. Method and Methodology
• Study site and period : The study was carried out in microbiology
laboratory, B and B Hospital from September 2010 to August 2011.
• Study population: A total of 1164 wound samples (swab, pus and
aspirates) were collected for culture and antibiotic sensitivity test was
performed. The patients with age of 1 year to 94 years were enrolled in the
study. The types of wound included were surgical wounds, trauma, burns,
ulcers, and other pyogenic wounds.
• Specimen collection: Wound pus and wound swab were collected on a
sterile swab or in a stoppered syringe without contamination. Two samples
were taken from each patient, one for culture and another for direct Gram
stain. The sample was taken to the laboratory for further processing
following standard microbiological laboratory procedures.
• Sample processing and isolation of organism:
 Macroscopic examination: Colour, odour and granules content were
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6. Method and Methodology
 Microscopic examination: The smear was gram stained and was
examined by using high dry (40X) and oil immersion (100X)
microscopy.
 Culture of specimen: The specimen was inoculated on Nutrient agar,
MacConkey agar and Blood agar and incubated at 37 Ċ for 24 to 48 hrs
aerobically.
 Identification of bacteria: Identification of the significant bacterial
isolates was done by using microbiological techniques as described in
the Bergey’s manual that involves colony characteristics, staining
reaction and various biochemical properties.
 Antimicrobial susceptibility testing: Antimicrobial Susceptibility
test was performed by Kirby-Bauer disk diffusion method following
Clinical and Laboratory Standard Institute guidelines (CLSI, 2010).
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7. Method and Methodology
Isolates showing resistance to two or more different classes of
antibiotic tested were considered as MDR.
 Preservation of the MDR isolates: MDR isolates in pure culture were
preserved in 20% glycerol containing Tryptic Soya broth and kept at -70°C
 Quality control: Accuracy of the overall procedure was monitored by
using S. aureus ATCC 25923 and E. coli ATCC 25922 as reference strain.
 Data analysis: Data were analysed by using WHONET 5.6.
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8. Results
Table 1: Age and Gender based distribution of patients with wound infection
Gender
Male Female
Age
Total Positive Prevalence Total Positive Prevalence
<15 58 29 50.0% 25 13 52.0%
15-30 317 151 47.6% 141 75 53.1%
30-45 282 146 51.7% 67 25 37.3%
>45 206 105 50.9% 68 29 42.6%
Total 863 431 49.9% 301 142 47.1%
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9. Results
Figure : Types and distribution of samples
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10. Results
Figure : Distribution of positive cases on season basis
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11. Results
Table : Types of bacteria
Bacterial isolates No. of isolate (N=573) Inpatients
(N=608)
Outpatients (N=556)
Gram positive Bacteria (N=146)
S. aureus 135 (92.46%) 87 48
CoNS 3 (2.05%) 2 1
β-haem. Streptococci 3 (2.05%) 2 1
Non haem. Streptococci 5 (3.42%) 3 2
Total 146 (25.48%)
Gram negative isolate (N=427)
E. coli
120(28.10%) 54 66
Pseudomonas spp. 132(30.91) 91 41
Klebsiella. Spp 41(9.60%) 28 13
Enterobacter spp. 60(14.05%) 36 24
P. mirabilis 3(0.70%) 3 0
P. vulgaris 2(0.46%) 0 2
Acinetobacter spp. 61(14.28%) 39 22
Citrobacter spp. 8(1.87%) 4 4
Total Kantipur 427 College 10/12/14 (74.52%)
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Total 573 349 (57.40%) 224 (40.29%)

12. Results
Table : Antibiotic sensitivity pattern of the bacterial isolates
Antibiotics Gram
positive
(N=146)
Gram negative
(N=427)
Total
(N=573)
S. aureus
(N=135)
Gentamycin 25 (17.12%) 214 (50.11%) 239 (41.71%) 19 (14.07%)
Amikacin 4 (2.73%) 146 (34.19%) 150 (26.17%) 4 (2.96%)
Ofloxcin 47 (32.19%) 251 (58.78%) 298 (52.00%) 38 (28.14%)
Ciprofloxacin 52 (36.61%) 260 (60.88%) 312 (54.45%) 44 (32.59%)
Amoxycillin 14 (9.58%) 357 (83.60%) 371 (64.74%) 13 (9.62%)
Cotrimoxazol 11 (7.53%) 240 (56.20%) 251 (43.80%) 10 (7.40%)
Chloramphenicol NT 260 (60.88%) ND NT
Erythromycin 48 (32.87%) NT ND 43 (31.85%)
Oxacillin 48 (32.87%) NT ND 46 (34.07%)
Penicillin NT NT ND 111 (82.22%)
Vancomycin NT NT ND 10 (7.40%)
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13. Conclusion
• Culture positivity in wound infection accounts about 50% of
the cases irrespective of their demographic status.
• Once infected it’s a problem for management as the etiological
agents isolated (S. aureus, E. coli, P. aurogenesa etc) are
resistant to most of commonly used antibiotics, so antibiotics
like Amikacin and Vancomycin (for S. aureus) can be used for
treatment and control.
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14. Recommendation
• Anaerobic bacteria also accounts in wound infection so, anaerobic
culture should be performed to establish exact prevalence of
bacterial wound infection.
• Proper disinfectants should be used while collecting wound
specimen to minimize commensals contamination.
• New antibiotics or high level antibiotics should be screened as
common antibiotics are usually seen to be resistant.
• The emergence of antibiotic resistance strains possess problem in
the management of wound infection. Thus, further molecular studies
should be recommended to reveal the real scenario.
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15. Acknowledgement
• Supervisors
• Faculty member Kantipur College of Medical Sciences
• Laboratory staff Kantipur College of Medical Sciences
• Staffs of Microbiology department, B and B hospital
• Family and friends
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